This folder contains codes and all datasets for ISME paper "Gut microbiota structure differs between honey bees in winter and summer":

1. In this paper we worked with 4 datasets:
		"0_df_mm.csv" - raw qPCR data for monthly sampled F/W bees (Fig1, S1-3)
		"0_df_pool.csv" - raw qPCR data for pooled F/N/W gut samples from 14 hives(Fig2)
		"0_df_exp.csv" - raw qPCR data for experimental bees (Fig3)
		"0_df_ind.csv" - raw qPCR data for individual F/N/W bees (Fig3)

2. These csv files were manually merged into "01_datasets.xlsx" file together with data for standard curves (sheet1)for absolute quantification calculations. The order of following sheets keeps the order above (sheets 2-5). This excel file serves as the main source of data for plots and statistics.

3. Code "01_rawData_calc.R" takes data from "01_datasets.xlsx" file and calculates for each target in sample:
	(i) target copy number (CopyNum)
	(ii) normalized copy number to the median actin value in the corresponding dataset (CopyNum_norm)
	(iii) number of bacterial cells (CellNum) based on known number of 16S rRNA gene loci in the genome of targeted bacterium
	(iv) number of normalised bacterial cells (CellNum_norm) calculated from CopyNum_norm in the same manner

	This code generates: 
		"02_df_mm_calc.csv"
		"02_df_pool_calc.csv"
		"02_df_exp_calc.csv"
		"02_df_ind_calc.csv"

4. 16S rRNA gene sequencing based community data requires four files for analysis:
	“03_df_sampleinfo.csv”: sample information will be used during DADA2 pipeline
	“03_df_sampleinfo2.csv”: sample information that will be used during downstream diversity and statistical analysis.
	“03_df_filt_16S.csv”: contains filtered community data with ASV sequences ontained at the end of DADA2 pipeline.
	“03_df_16S_phylo.csv”: contains community data after classifying ASVs into defined phylotypes.


5. Codes "2_functions.R" and "2_Dada2_Pipelines.R" serve to generate functions to be used during statistical anaysis, and to generate community data 16S rRNA gene amplicon sequences for downstream analysis. 

6. Codes "3_Plots_Figures_1_S1_S2_S3.R”, "3_Plots_Stats_Figures2_S4.R”, “3_Plots_Figure_3_S5.R”, and “3_Stats_Figures1_3_S5.R” serve to generate plots and perform stats on data.

FOR CERTAIN ANALYSIS THE '0'/'ZERO' VALUES SHOULD BE CHANGED TO '1' FOR LOG TRANSFORMATION.